Lettuce variety 81-90 RZ

ABSTRACT

The present invention relates to a  Lactuca sativa  seed designated 81-90 RZ, which exhibits a combination of traits including resistance to downy mildew ( Bremia lactucae ) races Bl:1-32 and Ca-I to Ca-VIII, resistance to currant-lettuce aphid ( Nasonovia ribisnigri ), resistance to lettuce mosaic virus (LMV), and resistance to corky root rot ( Sphingomonas suberifaciens ). The present invention also relates to a  Lactuca sativa  plant produced by growing the 81-90 RZ seed. The invention further relates to methods for producing the lettuce cultivar, represented by lettuce variety 81-90 RZ.

RELATED APPLICATIONS AND INCORPORATION BY REFERENCE

This application claims priority to U.S. provisional patent applicationSer. No. 62/250,078, filed Nov. 3, 2015.

The foregoing applications, and all documents cited therein or duringtheir prosecution (“appln cited documents”) and all documents cited orreferenced in the appln cited documents, and all documents cited orreferenced herein (“herein cited documents”), and all documents cited orreferenced in herein cited documents, together with any manufacturer'sinstructions, descriptions, product specifications, and product sheetsfor any products mentioned herein or in any document incorporated byreference herein, are hereby incorporated herein by reference, and maybe employed in the practice of the invention. More specifically, allreferenced documents are incorporated by reference to the same extent asif each individual document was specifically and individually indicatedto be incorporated by reference.

FIELD OF THE INVENTION

The present invention relates to a new lettuce (Lactuca sativa) varietyof the Batavia type which may exhibit a combination of traits includingresistance to downy mildew (Bremia lactucae) races Bl:1-32 and Ca-I toCa-VIII, resistance to currant-lettuce aphid (Nasonovia ribisnigri),resistance to lettuce mosaic virus (LMV), and resistance to corky rootrot (Sphingomonas suberifaciens).

BACKGROUND OF THE INVENTION

All cultivated forms of lettuce belong to the highly polymorphicspecies, Lactuca sativa, which is grown for its edible head and leaves.As a crop, lettuces are grown commercially wherever environmentalconditions permit the production of an economically viable yield.

Lactuca sativa is in the Cichoreae tribe of the Asteraceae (Compositae)family. Lettuce is related to chicory, sunflower, aster, scorzonera,dandelion, artichoke and chrysanthemum. Sativa is one of about 300species in the genus Lactuca.

Lettuce cultivars are susceptible to a number of pests and diseases suchas downy mildew (Bremia lactucae). Every year this disease leads tomillions of dollars of lost lettuce crop throughout the world. Downymildew (Bremia lactucae) is highly destructive on lettuce grown atrelatively low temperature and high humidity. Downy mildew is caused bya fungus, Bremia lactucae, which can be one of the following strains:NL1, NL2, NL4, NL5, NL6, NL7, NL10, NL12, NL13, NL14, NL15, NL16, Bl:17,Bl:18, Bl:20, Bl:21, Bl:22, Bl:23, Bl:24, Bl:25, Bl:26, Bl:27, Bl:28,Bl:29, Bl:30, Bl:31, Bl:32 (Van Ettekoven, K. et al., “Identificationand denomination of ‘new’ races of Bremia lactucae,” In: Lebeda, A. andKristkova, E (eds.), Eucarpia Leafy Vegetables, 1999, PalackyUniversity, Olomouc, Czech Republic, pp. 171-175; Van der Arend et al.“Identification and denomination of “new” races of Bremia lactucae inEurope by IBEB until 2002.” In: Van Hintum, Th et al. (eds.), EucarpiaLeafy Vegetables Conference 2003, Centre for Genetic Resources,Wageningen, The Netherlands, p. 151; Plantum NL (Dutch association forbreeding, tissue culture, production and trade of seeds and youngplants), Van der Arend et al. “Identification and denomination of “new”races of Bremia lactucae in Europe by IBEB until 2002.” In: Van Hintum,Th et al. (eds.), Eucarpia Leafy Vegetables Conference 2003, Centre forGenetic Resources, Wageningen, The Netherlands, p. 151; Plantum NL(Dutch association for breeding, tissue culture, production and trade ofseeds and young plants); IBEB press release “New race of Bremia lactucaeBl:27 identified and nominated”, May 2010; Plantum NL (Dutch associationfor breeding, tissue culture, production and trade of seeds and youngplants), “New race of Bremia lactucae Bl:28 identified and nominated”,March 2011; Plantum NL (Dutch association for breeding, tissue culture,production and trade of seeds and young plants), IBEB press release,“New races of Bremia lactucae, Bl:29, Bl:30 and Bl:31 identified andnominated”, August 2013; Plantum NL (Dutch association for breeding,tissue culture, production and trade of seeds and young plants), IBEBpress release, “A new race of Bremia lactucae, Bl:32 identified andnominated in Europe”, May 2015), Ca-I, Ca-IIA, Ca-IIB, Ca-III, Ca-IV(Schettini, T. M., Legg, E. J., Michelmore, R. W., 1991. Insensitivityto metalaxyl in California populations of Bremia lactucae and resistanceof California lettuce cultivars to downy mildew. Phytopathology 81(1).p. 64-70), and Ca-V, Ca-VI, Ca-VII, Ca-VIII (Michelmore R. & Ochoa. 0.“Breeding Crisphead Lettuce.” In: California Lettuce Research Board,Annual Report 2005-2006, 2006, Salinas, Calif., pp. 55-68).

Downy mildew causes pale, angular, yellow areas bounded by veins on theupper leaf surfaces. Sporulation occurs on the opposite surface of theleaves. The lesions eventually turn brown, and they may enlarge andcoalesce. These symptoms typically occur first on the lower leaves ofthe lettuce, but under ideal conditions may move into the upper leavesof the head. When the fungus progresses to this degree, the head cannotbe harvested. Less severe damage requires the removal of more leavesthan usual, especially when the lettuce reaches its final destination.

Of the various species of aphids that feed on lettuce, thecurrant-lettuce aphid (Nasonovia ribisnigri) is the most destructivespecies because it feeds both on the leaves of the lettuce as well asthe heart of the lettuce, making it difficult to control withconventional insecticides. The lettuce aphid feeds by sucking sap fromthe lettuce leaves. Although direct damage to the lettuce may belimited, its infestation has serious consequences because the presenceof aphids makes lettuce unacceptable to consumers.

Corky root rot is caused by a soil-borne bacterium, Sphingomonassuberifaciens, which is prevalent in most coastal lettuce growing areas.Corky root rot affects both leaf and head lettuce varieties. The diseasetypically is more severe when soil temperatures are warmer and in fieldswhere lettuce is grown consecutively. High soil nitrate levels alsoincrease disease severity. Early symptoms of corky root rot are yellowbands on tap and lateral roots of lettuce seedlings. The yellow areasgradually expand, taking on a greenish-brown color and developing cracksand rough areas on the surface of the root. As disease severityincreases, the entire tap root may become brown, severely cracked, andnonfunctional; the feeder root system also may be reduced and damaged.At this point, roots are very brittle and easily break off whenexamined. Corky root also may cause internal discoloration of the root.When the root is severely diseased, aboveground symptoms consist ofwilting during warm temperatures, stunting of plants, and general poorand uneven growth.

Lettuce mosaic virus (LMV) mainly infects lettuce seeds, which is theprimary way that the virus is introduced to lettuce in the fields, butalso can infect numerous crops and weeds, thereby creating reservoirs ofthe virus. LMV also can be vectored by aphids, which spread the viruswithin a lettuce field and introduce it into lettuce fields frominfected weeds and crops outside the field.

Symptoms of LMV vary greatly. Leaves of plants that are infected at ayoung stage are stunted, deformed and (in some varieties) show a mosaicor mottling pattern. Such plants rarely grow to full size; head lettucevarieties infected early fail to form heads. Plants that are infectedlater in the growth cycle show a different set of symptoms. These plantsmay reach full size, but the older outer leaves turn yellow, twisted,and otherwise are deformed. On head lettuce, the wrapper leaves oftenwill curve back away from the head and developing heads may be deformed.In some cases, brown necrotic flecks occur on the wrapper leaves.

The Leaf lettuce type is appreciated by consumers for its crisp,pleasant tasting leaves. In Europe, this type of lettuce is oftenreferred to as Batavia lettuce. Leaf lettuces grow upright, with looseleaves and are non-heading.

Citation or identification of any document in this application is not anadmission that such document is available as prior art to the presentinvention.

SUMMARY OF THE INVENTION

There exists a need, therefore, for a lettuce variety which exhibits acombination of traits including resistance to downy mildew (Bremialactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistance tocurrant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens).

The present invention addresses this need by providing a new type oflettuce (Lactuca sativa) variety, designated 81-90 RZ. Lettuce cultivar81-90 RZ exhibits a combination of traits including resistance to downymildew (Bremia lactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistanceto currant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens).

The present invention provides seeds of lettuce cultivar 81-90 RZ, whichhave been deposited with the National Collections of Industrial, Marineand Food Bacteria (NCIMB) in Bucksburn, Aberdeen AB21 9YA, Scotland, UKand have been assigned NCIMB Accession No. 42472.

In one embodiment, the invention provides a lettuce plant designated81-90 RZ, representative seed of which have been deposited under NCIMBAccession No. 42472, wherein said lettuce plant may comprise acombination of traits including resistance to downy mildew (Bremialactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistance tocurrant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens).

In one embodiment, the invention provides a lettuce plant designated81-90 RZ which may exhibit a combination of traits including resistanceto downy mildew (Bremia lactucae) races Bl:1-32 and Ca-I to Ca-VIII,resistance to currant-lettuce aphid (Nasonovia ribisnigri), resistanceto lettuce mosaic virus (LMV), resistance to corky root rot(Sphingomonas suberifaciens) and intense anthocyanin concentration ofthe mature leaves, representative seed of which have been depositedunder NCIMB Accession No. 42472.

In one embodiment, the invention provides a lettuce plant designated81-90 RZ, representative seed of which have been deposited under NCIMBAccession No. 42472.

In one embodiment, the invention provides for a lettuce plant which maycomprise genetic information for so exhibiting a combination of traitsincluding resistance to downy mildew (Bremia lactucae) races Bl:1-32 andCa-I to Ca-VIII, resistance to currant-lettuce aphid (Nasonoviaribisnigri), resistance to lettuce mosaic virus (LMV), and resistance tocorky root rot (Sphingomonas suberifaciens), wherein the geneticinformation is as contained in a plant, representative seed of whichhaving been deposited under NCIMB Accession No. 42472.

In one embodiment, the invention provides for a lettuce plant exhibitinga combination of traits including resistance to downy mildew (Bremialactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistance tocurrant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens), and having the genetic information for so exhibiting thecombination of traits, wherein the genetic information is as containedin a plant, representative seed of which having been deposited underNCIMB Accession No. 42472. In an embodiment of the present invention,there also is provided parts of a lettuce plant of the invention, whichmay include parts of a lettuce plant exhibiting a combination of traitsincluding resistance to downy mildew (Bremia lactucae) races Bl:1-32 andCa-I to Ca-VIII, resistance to currant-lettuce aphid (Nasonoviaribisnigri), resistance to lettuce mosaic virus (LMV), and resistance tocorky root rot (Sphingomonas suberifaciens), or parts of a lettuce planthaving any of the aforementioned resistance(s) and a combination oftraits including one or more morphological or physiologicalcharacteristics tabulated herein, including parts of lettuce variety81-90 RZ, wherein the plant parts are involved in sexual reproduction,which include, without limitation, microspores, pollen, ovaries, ovules,embryo sacs or egg cells and/or wherein the plant parts are suitable forvegetative reproduction, which include, without limitation, cuttings,roots, stems, cells or protoplasts and/or wherein the plant parts aretissue culture of regenerable cells in which the cells or protoplasts ofthe tissue culture are derived from a tissue such as, for example andwithout limitation, leaves, pollen, embryos, cotyledon, hypocotyls,meristematic cells, roots, root tips, anthers, flowers, seeds or stems.The plants of the invention from which such parts may come from includethose wherein representative seed of which has been deposited underNCIMB Accession No. 42472 or lettuce variety or cultivar designated81-90 RZ, as well as seed from such a plant, plant parts of such a plant(such as those mentioned herein) and plants from such seed and/orprogeny of such a plant, advantageously progeny exhibiting suchcombination of such traits, each of which, is within the scope of theinvention; and such combination of traits.

In another embodiment there is a plant grown from seeds, representativeseed of which having been deposited under NCIMB Accession No. 42472.

In a further embodiment there is a plant regenerated from theabove-described plant parts or regenerated from the above-describedtissue culture. Advantageously such a plant may have morphologicaland/or physiological characteristics of lettuce variety 81-90 RZ and/orof a plant grown from seed, representative seed of which having beendeposited under NCIMB Accession No. 42472—including without limitationsuch plants having all of the morphological and physiologicalcharacteristics of lettuce variety 81-90 RZ and/or of plant grown fromseed, representative seed of which having been deposited under NCIMBAccession No. 42472. Advantageously, such a plant demonstrates thetraits of resistance to downy mildew (Bremia lactucae) races Bl:1-32 andCa-I to Ca-VIII, resistance to currant-lettuce aphid (Nasonoviaribisnigri), resistance to lettuce mosaic virus (LMV), and resistance tocorky root rot (Sphingomonas suberifaciens).

Accordingly, in still a further embodiment, there is provided a lettuceplant having all of the morphological and physiological characteristicsof lettuce variety 81-90 RZ, representative seed of which having beendeposited under NCIMB Accession No. 42472. Such a plant may be grownfrom the seeds, regenerated from the above-described plant parts, orregenerated from the above-described tissue culture. A lettuce planthaving any of the aforementioned resistance(s), and one or moremorphological or physiological characteristics recited or tabulatedherein, and a lettuce plant advantageously having all of theaforementioned resistances and the characteristics recited and tabulatedherein, are preferred. Parts of such plants—such as those plant partsabove-mentioned—are encompassed by the invention.

In one embodiment, there is provided a method for producing a progeny oflettuce cultivar 81-90 RZ which may comprise crossing the a plantdesignated 81-90 RZ with itself or with another lettuce plant,harvesting the resultant seed, and growing said seed.

In a further embodiment there is provided a progeny plant produced bysexual or vegetative reproduction, grown from seeds, regenerated fromthe above-described plant parts, or regenerated from the above-describedtissue culture of the lettuce cultivar or a progeny plant thereof,representative seed of which having been deposited under NCIMB AccessionNo. 42472. The progeny may have any of the aforementioned resistance(s),and one or more morphological or physiological characteristics recitedor tabulated herein, and a progeny plant advantageously having all ofthe aforementioned resistances and the characteristics recited andtabulated herein, are preferred. Advantageously, the progeny demonstratethe traits of resistance to downy mildew (Bremia lactucae) races Bl:1-32and Ca-I to Ca-VIII, resistance to currant-lettuce aphid (Nasonoviaribisnigri), resistance to lettuce mosaic virus (LMV), and resistance tocorky root rot (Sphingomonas suberifaciens).

Progeny of the lettuce variety 81-90 RZ may be modified in one or moreother characteristics, in which the modification is a result of, forexample and without limitation, mutagenesis or transformation with atransgene.

In still another embodiment, the present invention provides progeny oflettuce cultivar 81-90 RZ produced by sexual or vegetative reproduction,grown from seeds, regenerated from the above-described plant parts, orregenerated from the above-described tissue culture of the lettucecultivar or a progeny plant thereof, in which the regenerated plantshows a combination of traits including resistance to downy mildew(Bremia lactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistance tocurrant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens).

In still a further embodiment, the invention provides a method ofproducing a hybrid lettuce seed which may comprise crossing a firstparent lettuce plant with a second parent lettuce plant and harvestingthe resultant hybrid lettuce seed, in which the first parent lettuceplant or the second parent lettuce plant may be a lettuce plant of theinvention, e.g. a lettuce plant having a combination of traits includingresistance to downy mildew (Bremia lactucae) races Bl:1-32 and Ca-I toCa-VIII, resistance to currant-lettuce aphid (Nasonovia ribisnigri),resistance to lettuce mosaic virus (LMV), and resistance to corky rootrot (Sphingomonas suberifaciens) and one or more morphological orphysiological characteristics tabulated herein, including a lettuceplant of lettuce cultivar 81-90 RZ, representative seed of which havingbeen deposited under 42472.

In another embodiment, the invention provides producing a lettuce plantwhich may exhibit a combination of traits including resistance to downymildew (Bremia lactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistanceto currant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens) which may comprise: crossing a mother lettuce plant witha father lettuce plant to produce a hybrid seed; growing said hybridseed to produce a hybrid plant; selfing said hybrid plant to produce F2progeny seed; growing said F2 progeny seed to produce F2-plants;selecting said F2-plants for exhibiting a combination of traitsincluding resistance to downy mildew (Bremia lactucae) races Bl:1-32 andCa-I to Ca-VIII, resistance to currant-lettuce aphid (Nasonoviaribisnigri), resistance to lettuce mosaic virus (LMV), and resistance tocorky root rot (Sphingomonas suberifaciens).

Advantageously the selfing and selection may be repeated; for example atleast once, or at least twice, thrice, four times, five times, six timesor more, to produce F3 or F4 or F5 or F6 or subsequent progeny,especially as progeny from F2 may exhibit the aforementioned combinationof traits, and may be desirable.

In still a further embodiment, the invention provides a method ofproducing a lettuce cultivar which may exhibit a combination of traitsincluding resistance to downy mildew (Bremia lactucae) races Bl:1-32 andCa-I to Ca-VIII, resistance to currant-lettuce aphid (Nasonoviaribisnigri), resistance to lettuce mosaic virus (LMV), and resistance tocorky root rot (Sphingomonas suberifaciens).

The invention even further relates to a method of producing lettucewhich may comprise: (a) cultivating to the vegetative plant stage aplant of lettuce variety 81-90 RZ, representative seed of which havingbeen deposited under NCIMB Accession No. 42472, and (b) harvestinglettuce leaves or heads from the plant. The invention furthercomprehends packaging and/or processing the lettuce plants, heads orleaves.

Accordingly, it is an object of the invention to not encompass withinthe invention any previously known product, process of making theproduct, or method of using the product such that Applicants reserve theright and hereby disclose a disclaimer of any previously known product,process, or method. It is further noted that the invention does notintend to encompass within the scope of the invention any product,process, or making of the product or method of using the product, whichdoes not meet the written description and enablement requirements of theUSPTO (35 U.S.C. § 112, first paragraph) or the EPO (Article 83 of theEPC), such that Applicants reserve the right and hereby disclose adisclaimer of any previously described product, process of making theproduct, or method of using the product.

It is noted that in this disclosure and particularly in the claims,terms such as “comprises”, “comprised”, and “comprising” and the like(e.g., “includes”, “included”, “including”, “contains”, “contained”,“containing”, “has”, “had”, “having”, etc.) can have the meaningascribed to them in US Patent law, i.e., they are open ended terms. Forexample, any method that “comprises,” “has” or “includes” one or moresteps is not limited to possessing only those one or more steps and alsocovers other unlisted steps. Similarly, any plant that “comprises,”“has” or “includes” one or more traits is not limited to possessing onlythose one or more traits and covers other unlisted traits. Similarly,the terms “consists essentially of” and “consisting essentially of” havethe meaning ascribed to them in US Patent law, e.g., they allow forelements not explicitly recited, but exclude elements that are found inthe prior art or that affect a basic or novel characteristic of theinvention. See also MPEP § 2111.03. In addition, the term “about” isused to indicate that a value includes the standard deviation of errorfor the device or method being employed to determine the value.

These and other embodiments are disclosed or are obvious from andencompassed by the following Detailed Description.

DEPOSIT

The Deposit with NCIMB Ltd, Ferguson Building, Craibstone Estate,Bucksburn, Aberdeen AB21 9YA, UK, on Oct. 23, 2015, under depositaccession number 42472 was made pursuant to the terms of the BudapestTreaty. Upon issuance of a patent, all restrictions upon the depositwill be removed, and the deposit is intended to meet the requirements of37 CFR § 1.801-1.809. The deposit will be irrevocably and withoutrestriction or condition released to the public upon the issuance of apatent and for the enforceable life of the patent. The deposit will bemaintained in the depository for a period of 30 years, or 5 years afterthe last request, or for the effective life of the patent, whichever islonger, and will be replaced if necessary during that period.

BRIEF DESCRIPTION OF THE DRAWINGS

The patent or application file contains at least one drawing executed incolor. Copies of this patent or patent application publication withcolor drawing(s) will be provided by the Office upon request and paymentof the necessary fee.

The following detailed description, given by way of example, but notintended to limit the invention solely to the specific embodimentsdescribed, may best be understood in conjunction with the accompanyingdrawing, in which:

FIG. 1 is an illustration of six different shapes of the fourth leaffrom a 20-day old seedling grown under optimal conditions.

FIG. 2 is a picture of the shape of the fourth leaf of 81-90 RZ.

FIG. 3 is a picture of the shape of a mature leaf of 81-90 RZ.

DETAILED DESCRIPTION OF THE INVENTION

The invention provides methods and compositions relating to plants,seeds and derivatives of a new lettuce variety herein referred to aslettuce variety 81-90 RZ. Lettuce variety 81-90 RZ is a uniform andstable line, distinct from other such lines.

In a preferred embodiment, the specific type of breeding method employedfor developing a lettuce cultivar is pedigree selection, where bothsingle plant selection and mass selection practices are employed.Pedigree selection, also known as the “Vilmorin system of selection,” isdescribed in Fehr, W., Principles of Cultivar Development, Volume I,MacMillan Publishing Co., which is hereby incorporated by reference.

When pedigree selection is applied, in general selection is firstpracticed among F₂ plants. In the next season, the most desirable F₃lines are first identified, and then desirable F₃ plants within eachline are selected. The following season and in all subsequentgenerations of inbreeding, the most desirable families are identifiedfirst, then desirable lines within the selected families are chosen, andfinally desirable plants within selected lines are harvestedindividually. A family refers to lines that were derived from plantsselected from the same progeny from the preceding generation.

Using this pedigree method, two parents may be crossed using anemasculated female and a pollen donor (male) to produce F₁ offspring.Lettuce is an obligate self-pollination species, which means that pollenis shed before stigma emergence, assuring 100% self-fertilization.Therefore, in order to optimize crossing, a method of misting may beused to wash the pollen off prior to fertilization to assure crossing orhybridization.

Parental varieties are selected from commercial varieties thatindividually exhibit one or more desired phenotypes. Additionally, anybreeding method involving selection of plants for the desired phenotypemay be used in the method of the present invention.

The F₁ may be self-pollinated to produce a segregating F₂ generation.Individual plants may then be selected which represent the desiredphenotype in each generation (F₃, F₄, F₅, etc.) until the traits arehomozygous or fixed within a breeding population.

A detailed description of the development of lettuce variety 81-90 RZ isdescribed in Table 1. The seed lot in year 9, seed lot 14R.4489, wasdeposited with the NCIMB under deposit number 42472.

TABLE 1 Year Description Location 0 Final F1-cross internal breedingline ‘35638’ × Aramon, France internal breeding line ‘19167’ inglasshouse. 1 F1 plant grown for F2 seed production in Aramon, Franceglasshouse. 2 F2 plant 07A.28813 selected in open Aramon, Francefieldfollowed by F3 seed 08A.48535 3 F3 plant 08A.26600 selected in openfield, Aramon, France followed by F4 seed 09A.32088 4 F4 plant 09A.70987selected in open field, Fijnaart, followed by F5 seed 10A.32078 theNetherlands 5 F5 plant 10A.71950 selected in open field, Fijnaart,followed by F6 seed 11A.31351 the Netherlands 6 F6 plant 11A.24664selected in open field, Le Thor, France followed by F7 seed 12A.31100 7F7 plant 12A.28775 selected in open field, Aramon, France followed by F8seed 13A.30374 8 F8 plant 13A.71471 selected in open field, Fijnaart,followed by F9 seed 13A.35775 the Netherlands 9 F9 line 13A.35775established uniform, Aramon, France multiplied in plastic tunnel: seedlot 14R.4489.

In one embodiment, a plant of the invention has all the morphologicaland physiological characteristics of lettuce variety 81-90 RZ. Thesecharacteristics of a lettuce plant of the invention, e.g. variety 81-90RZ, are summarized in Tables 2 and 3.

Next to the morphological and physiological characteristics mentioned inTables 2 and 3, a plant of the invention also exhibits resistance todowny mildew (Bremia lactucae Regel.).

As used herein resistance against Bremia lactucae is defined as theability of a plant to resist infection by each of the various strainsBl:1 to Bl:32, Ca-I to Ca-VIII of Bremia lactucae Regel. in all stagesbetween the seedling stage and the harvestable plant stage. Bl:1 toBl:32 means strains NL1, NL2, NL4, NL5, NL6, NL7, NL10, NL12, NL13,NL14, NL15, NL16, Bl:17, Bl:18, Bl:20, Bl:21, Bl:22, Bl:23, Bl:24,Bl:25, Bl:26, Bl:27, Bl:28, Bl:29, Bl:30, Bl:31, Bl:32 (Van Ettekoven K,Van der Arend A J M, 1999. identification and denomination of ‘new’races of Bremia lactucae. In: Lebeda A, Kristkova E (eds.) Eucarpialeafy vegetables '99. Palacky University, Olomouc, Czech Republic, 1999:171-175; Van der Arend, A. J. M., Gautier, J., Guenard, M., Michel, H.,Moreau, B., de Ruijter, J., Schut, J. W. and de Witte, I. (2003).Identification and denomination of ‘new’ races of Bremia lactucae inEurope by IBEB until 2002. In: Eucarpia leafy vegetables 2003.Proceedings of the Eucarpia Meeting on leafy vegetables genetics andbreeding. Noorwijkerhout, The Netherlands. Eds. Van Hintum T., LebedaA., Pink D., Schut J. pp 151-160; Van der Arend A J M, Gautier J,Grimault V, Kraan P, Van der Laan R, Mazet J, Michel H, Schut J W,Smilde D, De Witte I (2006) Identification and denomination of “new”races of Bremia lactucae in Europe by IBEB until 2006; incorporatedherein by reference; Plantum NL (Dutch association for breeding, tissueculture, production and trade of seeds and young plants), IBEB pressrelease, “New race of Bremia lactucae Bl:27 identified and nominated”,May 2010; Plantum NL (Dutch association for breeding, tissue culture,production and trade of seeds and young plants), IBEB press release,“New race of Bremia lactucae Bl:28 identified and nominated”, March2011; Plantum NL (Dutch association for breeding, tissue culture,production and trade of seeds and young plants), IBEB press release,“New races of Bremia lactucae, Bl:29, Bl:30 and Bl:31 identified andnominated”, August 2013; Plantum NL (Dutch association for breeding,tissue culture, production and trade of seeds and young plants), IBEBpress release, “A new race of Bremia lactucae, Bl:32 identified andnominated in Europe”, May 2015). Ca-I to Ca-VIII means Ca-I, Ca-IIA,Ca-IIB, Ca-III, Ca-IV (Schettini, T. M., Legg, E. J., Michelmore, R. W.,1991. Insensitivity to metalaxyl in California populations of Bremialactucae and resistance of California lettuce cultivars to downy mildew,Phytopathology 81(1). p. 64-70), and Ca-V, Ca-VI, Ca-VII, Ca-VIII(Michelmore R. & Ochoa. O. “Breeding Crisphead Lettuce.” In: CaliforniaLettuce Research Board, Annual Report 2005-2006, 2006, Salinas, Calif.,pp. 55-68).

Resistance typically is tested by two interchangeable methods, describedby Bonnier, F. J. M. et al. (Euphytica, 61(3):203-211, 1992;incorporated herein by reference). One method involves inoculating 7-dayold seedlings, and observing sporulation 10 to 14 days later. The othermethod involves inoculating leaf discs with a diameter of 18 mm obtainedfrom a non-senescent, fully grown true leaf and observing sporulation 10days later.

As used herein, resistance against Nasonovia ribisnigri (Mosley), orcurrant-lettuce aphid, is defined as the plant characteristic whichresults in a non-feeding response of the aphid on the leaves of theplant in all stages between 5 true-leaf stage and harvestable plantstage (U.S. Pat. No. 5,977,443 to Jansen, J. P. A., “Aphid Resistance inComposites,” p. 12, 1999; incorporated herein by reference).

Resistance is tested by spreading at least ten aphids of biotype Nr:0 ona plant in a plant stage between 5 true leaves and harvestable stage,and observing the density of the aphid population on the plant as wellas the growth reduction after 14 days in a greenhouse, with temperaturesettings of 23 degrees Celsius in daytime and 21 degrees Celsius atnight. Day length is kept at 18 hours by assimilation lights.

As used herein, resistance against lettuce mosaic virus (LMV) may bedefined as the ability of the plant to grow normally after LMV infectionand to inhibit the virus transmission to seed. Resistance may be testedby mechanical inoculation of young plants in a climate cell orgreenhouse, as described by Pink, D. A. C. et al. (Plant Pathology,41(1): 5-12, 1992), incorporated herein by reference. Inoculatedresistant plants grow just as well as uninoculated plants and show nochlorosis or mosaic symptoms. The LMV isolate, which may be used fortesting, is Ls-1 (International Union for the Protection of NewVarieties of Plants [UPOV], Guidelines for the conduct of tests fordistinctness, uniformity and stability; 30 lettuce (Lactuca sativa L.),2002, p. 35; incorporated herein by reference).

As used herein, resistance against Sphingomonas suberifaciens, alsoknown as corky root rot or Rhizomonas suberifaciens, is defined as theability of the plant to grow relatively normally in a corkyroot-infected soil (Yabuuchi, E, et al., Microbiol. Immunol., 43;339-349, 1997). As the resistance of the lettuce plant to the bacteriumis incomplete, a susceptible and a resistant standard variety, forexample “Parris Island” (susceptible) and “Tall Guzmaine” (resistant),should be used for comparison. Resistance is tested with a young plantin a greenhouse using the CA1-strain of the bacterium, as described byBrown, P. R. et al. (Phytopathology, 78:1145-1150, 1988), incorporatedherein by reference.

As used herein, a plant with a non-heading plant habit is a plant whichdoes not make a head, i.e. with no head formation (International Unionfor the Protection of New Varieties of Plants [UPOV]), Guidelines forthe conduct of tests for distinctness, uniformity and stability; lettuce(Lactuca sativa L.), 2002; incorporated herein by reference). As areference varieties Ruby and Louxal can be used. It is scorednon-heading for head shape in Table 3.

As used herein, intensely red leaves are defined as leaves with ananthocyanin distribution throughout the leaf and an anthocyaninconcentration which is equal in comparison to the variety Cherokee. Theleaves of 81-90 RZ have an anthocyanin distribution throughout and anintense anthocyanin concentration, comparable to that of the leaves ofCherokee which also has an anthocyanin distribution throughout, and anintense anthocyanin concentration.

In Table 2, the seed color, cotyledon shape and characteristics of thefourth leaf of “81-90 RZ” are compared with “Louxal” and “Ruby”.

TABLE 2 Character “81-90 RZ” Louxal Ruby Plant Type Batavia BataviaCutting/leaf Seed Color White White White Cotyledon Shape IntermediateIntermediate Intermediate Shape of Fourth Leaf Elongated Oval ElongatedRolling of Fourth Leaf Present Present Present Cupping of Fourth LeafAbsent Absent Absent Fourth Leaf Entire Entire Entire Apical MarginFourth Leaf Coarsely Coarsely Coarsely Basal Margin dentate dentatedentate Undulation Slight Slight Slight Leaf color Red Red Light greenAnthocyanin distribution Throughout Throughout Throughout Anthocyaninconcentration Moderate Moderate Light to light

In Table 3, the mature leaf and head characteristics of “81-90 RZ” arecompared with “Louxal” and “Ruby”. RHS=Royal Horticultural Society colorchart code.

TABLE 3 Character “81-90 RZ” Louxal Ruby Leaf Color Red Red RedAnthocyanin Throughout Throughout Throughout Distribution AnthocyaninIntense Moderate Moderate concentration Margin Incision Absent shallowAbsent shallow Absent shallow Depth Margin Shallowely ShallowelyShallowely Indentation dentate dentate dentate Undulations of theModerate Moderate Moderate Apical Margin Leaf Size Medium Medium MediumLeaf Glossiness Glossy Glossy Glossy to moderate Leaf Blistering StrongStrong Strong Leaf Thickness Thin Thin Very thin Head Shape Non-headingNon-heading Non-heading Butt Shape Rounded Rounded Rounded MidribModerately Moderately Moderately raised raised raised

In an embodiment, the invention relates to lettuce plants that has allthe morphological and physiological characteristics of the invention andhave acquired said characteristics by introduction of the geneticinformation that is responsible for the characteristics from a suitablesource, either by conventional breeding, or genetic modification, inparticular by cisgenesis or transgenesis. Cisgenesis is geneticmodification of plants with a natural gene, coding for an (agricultural)trait, from the crop plant itself or from a sexually compatible donorplant. Transgenesis is genetic modification of a plant with a gene froma non-crossable species or a synthetic gene.

Just as useful traits that may be introduced by backcrossing, usefultraits may be introduced directly into the plant of the invention, beinga plant of lettuce variety 81-90 RZ, by genetic transformationtechniques; and, such plants of lettuce variety 81-90 RZ that haveadditional genetic information introduced into the genome or thatexpress additional traits by having the DNA coding there for introducedinto the genome via transformation techniques, are within the ambit ofthe invention, as well as uses of such plants, and the making of suchplants.

Genetic transformation may therefore be used to insert a selectedtransgene into the plant of the invention, being a plant of lettucevariety 81-90 RZ or may, alternatively, be used for the preparation oftransgenes which may be introduced by backcrossing. Methods for thetransformation of plants, including lettuce, are well known to those ofskill in the art.

Vectors used for the transformation of lettuce cells are not limited solong as the vector may express an inserted DNA in the cells. Forexample, vectors which may comprise promoters for constitutive geneexpression in lettuce cells (e.g., cauliflower mosaic virus 35Spromoter) and promoters inducible by exogenous stimuli may be used.Examples of suitable vectors include pBI binary vector. The “lettucecell” into which the vector is to be introduced includes various formsof lettuce cells, such as cultured cell suspensions, protoplasts, leafsections, and callus. A vector may be introduced into lettuce cells byknown methods, such as the polyethylene glycol method, polycationmethod, electroporation, Agrobacterium-mediated transfer, particlebombardment and direct DNA uptake by protoplasts. To effecttransformation by electroporation, one may employ either friabletissues, such as a suspension culture of cells or embryogenic callus oralternatively one may transform immature embryos or other organizedtissue directly. In this technique, one would partially degrade the cellwalls of the chosen cells by exposing them to pectin-degrading enzymes(pectolyases) or mechanically wound tissues in a controlled manner.

A particularly efficient method for delivering transforming DNA segmentsto plant cells is microprojectile bombardment. In this method, particlesare coated with nucleic acids and delivered into cells by a propellingforce. Exemplary particles include those which may be comprised oftungsten, platinum, and preferably, gold. For the bombardment, cells insuspension are concentrated on filters or solid culture medium.Alternatively, immature embryos or other target cells may be arranged onsolid culture medium. The cells to be bombarded are positioned at anappropriate distance below the macroprojectile stopping plate. Anillustrative embodiment of a method for delivering DNA into plant cellsby acceleration is the Biolistics Particle Delivery System, which may beused to propel particles coated with DNA or cells through a screen, suchas a stainless steel or Nytex screen, onto a surface covered with targetlettuce cells. The screen disperses the particles so that they are notdelivered to the recipient cells in large aggregates. It is believedthat a screen intervening between the projectile apparatus and the cellsto be bombarded reduces the size of projectiles aggregate and maycontribute to a higher frequency of transformation by reducing thedamage inflicted on the recipient cells by projectiles that are toolarge. Microprojectile bombardment techniques are widely applicable, andmay be used to transform virtually any plant species, including a plantof lettuce variety 81-90 RZ.

Agrobacterium-mediated transfer is another widely applicable system forintroducing gene loci into plant cells. An advantage of the technique isthat DNA may be introduced into whole plant tissues, thereby bypassingthe need for regeneration of an intact plant from a protoplast.Agrobacterium transformation vectors are capable of replication in E.coli as well as Agrobacterium, allowing for convenient manipulations.Moreover, advances in vectors for Agrobacterium-mediated gene transferhave improved the arrangement of genes and restriction sites in thevectors to facilitate the construction of vectors capable of expressingvarious polypeptide coding genes. The vectors have convenientmulti-linker regions flanked by a promoter and a polyadenylation sitefor direct expression of inserted polypeptide coding genes.Additionally, Agrobacterium containing both armed and disarmed Ti genesmay be used for transformation. In those plant strains whereAgrobacterium-mediated transformation is efficient, it is the method ofchoice because of the facile and defined nature of the gene locustransfer. The use of Agrobacterium-mediated plant integrating vectors tointroduce DNA into plant cells, including lettuce plant cells, is wellknown in the art (See, e.g., U.S. Pat. Nos. 7,250,560 and 5,563,055).

Transformation of plant protoplasts also may be achieved using methodsbased on calcium phosphate precipitation, polyethylene glycol treatment,electroporation, and combinations of these treatments.

A number of promoters have utility for plant gene expression for anygene of interest including but not limited to selectable markers,scoreable markers, genes for pest tolerance, disease resistance,nutritional enhancements and any other gene of agronomic interest.Examples of constitutive promoters useful for lettuce plant geneexpression include, but are not limited to, the cauliflower mosaic virus(CaMV) P-35S promoter, a tandemly duplicated version of the CaMV 35Spromoter, the enhanced 35S promoter (P-e35S), the nopaline synthasepromoter, the octopine synthase promoter, the figwort mosaic virus(P-FMV) promoter (see U.S. Pat. No. 5,378,619), an enhanced version ofthe FMV promoter (P-eFMV) where the promoter sequence of P-FMV isduplicated in tandem, the cauliflower mosaic virus 19S promoter, asugarcane bacilliform virus promoter, a commelina yellow mottle viruspromoter, the promoter for the thylakoid membrane proteins from lettuce(psaD, psaF, psaE, PC, FNR, atpC, atpD, cab, rbcS) (see U.S. Pat. No.7,161,061), the CAB-1 promoter from lettuce (see U.S. Pat. No.7,663,027), the promoter from maize prolamin seed storage protein (seeU.S. Pat. No. 7,119,255), and other plant DNA virus promoters known toexpress in plant cells. A variety of plant gene promoters that areregulated in response to environmental, hormonal, chemical, and/ordevelopmental signals may be used for expression of an operably linkedgene in plant cells, including promoters regulated by (1) heat, (2)light (e.g., pea rbcS-3A promoter, maize rbcS promoter, or chlorophylla/b-binding protein promoter), (3) hormones, such as abscisic acid, (4)wounding (e.g., wunl, or (5) chemicals such as methyl jasmonate,salicylic acid, or Safener. It may also be advantageous to employorgan-specific promoters.

Exemplary nucleic acids which may be introduced to the lettuce varietyof this invention include, for example, DNA sequences or genes fromanother species, or even genes or sequences which originate with or arepresent in lettuce species, but are incorporated into recipient cells bygenetic engineering methods rather than classical reproduction orbreeding techniques. However, the term “exogenous” is also intended torefer to genes that are not normally present in the cell beingtransformed, or perhaps simply not present in the form, structure, etc.,as found in the transforming DNA segment or gene, or genes which arenormally present and that one desires to express in a manner thatdiffers from the natural expression pattern, e.g., to over-express.Thus, the term “exogenous” gene or DNA is intended to refer to any geneor DNA segment that is introduced into a recipient cell, regardless ofwhether a similar gene may already be present in such a cell. The typeof DNA included in the exogenous DNA may include DNA which is alreadypresent in the plant cell, DNA from another plant, DNA from a differentorganism, or a DNA generated externally, such as a DNA sequencecontaining an antisense message of a gene, or a DNA sequence encoding asynthetic or modified version of a gene.

Many hundreds if not thousands of different genes are known and couldpotentially be introduced into a plant of lettuce variety 81-90 RZ.Non-limiting examples of particular genes and corresponding phenotypesone may choose to introduce into a lettuce plant include one or moregenes for insect tolerance, pest tolerance such as genes for fungaldisease control, herbicide tolerance, and genes for quality improvementssuch as yield, nutritional enhancements, environmental or stresstolerances, or any desirable changes in plant physiology, growth,development, morphology or plant product(s).

Alternatively, the DNA coding sequences may affect these phenotypes byencoding a non-translatable RNA molecule that causes the targetedinhibition of expression of an endogenous gene, for example viaantisense- or cosuppression-mediated mechanisms. The RNA could also be acatalytic RNA molecule (i.e., a ribozyme) engineered to cleave a desiredendogenous mRNA product. Thus, any gene which produces a protein or mRNAwhich expresses a phenotype or morphology change of interest is usefulfor the practice of the present invention. (See also U.S. Pat. No.7,576,262, “Modified gene-silencing RNA and uses thereof.”)

U.S. Pat. Nos. 7,230,158, 7,122,720, 7,081,363, 6,734,341, 6,503,732,6,392,121, 6,087,560, 5,981,181, 5,977,060, 5,608,146, 5,516,667, eachof which, and all documents cited therein are hereby incorporated hereinby reference, consistent with the above INCORPORATION BY REFERENCEsection, are additionally cited as examples of U.S. Patents that mayconcern transformed lettuce and/or methods of transforming lettuce orlettuce plant cells, and techniques from these US patents, as well aspromoters, vectors, etc., may be employed in the practice of thisinvention to introduce exogenous nucleic acid sequence(s) into a plantof lettuce variety 81-90 RZ (or cells thereof), and exemplify someexogenous nucleic acid sequence(s) which may be introduced into a plantof lettuce variety 81-90 RZ (or cells thereof) of the invention, as wellas techniques, promoters, vectors etc., to thereby obtain further plantsof lettuce variety 81-90 RZ, plant parts and cells, seeds, otherpropagation material harvestable parts of these plants, etc. of theinvention, e.g. tissue culture, including a cell or protoplast, such asan embryo, meristem, cotyledon, pollen, leaf, anther, root, root tip,pistil, flower, seed or stalk.

The invention further relates to propagation material for producingplants of the invention. Such propagation material may comprise interalia seeds of the claimed plant and parts of the plant that are involvedin sexual reproduction. Such parts are for example selected from thegroup consisting of seeds, microspores, pollen, ovaries, ovules, embryosacs and egg cells. In addition, the invention relates to propagationmaterial which may comprise parts of the plant that are suitable forvegetative reproduction, for example cuttings, roots, stems, cells,protoplasts.

According to a further aspect thereof the propagation material of theinvention may comprise a tissue culture of the claimed plant. The tissueculture may comprise regenerable cells. Such tissue culture may bederived from leaves, pollen, embryos, cotyledon, hypocotyls,meristematic cells, roots, root tips, anthers, flowers, seeds and stems.(See generally U.S. Pat. No. 7,041,876 on lettuce being recognized as aplant that may be regenerated from cultured cells or tissue).

Also, the invention comprehends methods for producing a seed of a “81-90RZ”-derived lettuce plant which may comprise (a) crossing a plant oflettuce variety 81-90 RZ, representative seed of which having beendeposited under NCIMB Accession No. 42472, with a second lettuce plant,and (b) whereby seed of a 81-90 RZ-derived lettuce plant form. Such amethod may further comprise (c) crossing a plant grown from 81-90RZ-derived lettuce seed with itself or with a second lettuce plant toyield additional 81-90 RZ-derived lettuce seed, (d) growing theadditional 81-90 RZ-derived lettuce seed of step (c) to yield additional81-90 RZ-derived lettuce plants, and (e) repeating the crossing andgrowing of steps (c) and (d) for an additional 3-10 generations tofurther generate 81-90 RZ-derived lettuce plants.

The invention further relates to the above methods that may furthercomprise selecting at steps b), d), and e), a 81-90 RZ-derived lettuceplant, exhibiting a combination of traits including resistance to downymildew (Bremia lactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistanceto currant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens).

In particular, the invention relates to methods for producing a seed ofa 81-90 RZ-derived lettuce plant which may comprise (a) crossing a plantof lettuce variety 81-90 RZ, representative seed of which having beendeposited under NCIMB Accession No. 42472, with a second lettuce plantand (b) whereby seed of a 81-90 RZ-derived lettuce plant forms, whereinsuch a method may further comprise (c) crossing a plant grown from 81-90RZ-derived lettuce seed with itself or with a second lettuce plant toyield additional 81-90 RZ-derived lettuce seed, (d) growing theadditional 81-90 RZ-derived lettuce seed of step (c) to yield additional81-90 RZ-derived lettuce plants and selecting plants exhibiting acombination of the traits including resistance to downy mildew (Bremialactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistance tocurrant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens), and (e) repeating the crossing and growing of steps (c)and (d) for an additional 3-10 generations to further generate 81-90RZ-derived lettuce plants that exhibit a combination of traits includingresistance to downy mildew (Bremia lactucae) races Bl:1-32 and Ca-I toCa-VIII, resistance to currant-lettuce aphid (Nasonovia ribisnigri),resistance to lettuce mosaic virus (LMV), and resistance to corky rootrot (Sphingomonas suberifaciens).

The invention additionally provides a method of introducing a desiredtrait into a plant of lettuce variety 81-90 RZ which may comprise: (a)crossing a plant of lettuce variety 81-90 RZ, representative seed ofwhich having been deposited under NCIMB Accession No. 42472, with asecond lettuce plant that may comprise a desired trait to produce F1progeny; (b) selecting an F1 progeny that may comprise the desiredtrait; (c) crossing the selected F1 progeny with a plant of lettucevariety 81-90 RZ, to produce backcross progeny; (d) selecting backcrossprogeny which may comprise the desired trait and the physiological andmorphological characteristic of a plant of lettuce variety 81-90 RZ;and, optionally, (e) repeating steps (c) and (d) one or more times insuccession to produce selected fourth or higher backcross progeny thatmay comprise the desired trait and all of the physiological andmorphological characteristics of a plant of lettuce variety 81-90 RZ,when grown in the same environmental conditions. The invention, ofcourse, includes a lettuce plant produced by this method.

Backcrossing may also be used to improve an inbred plant. Backcrossingtransfers a specific desirable trait from one inbred or non-inbredsource to an inbred that lacks that trait. This may be accomplished, forexample, by first crossing a superior inbred (A) (recurrent parent) to adonor inbred (non-recurrent parent), which carries the appropriate locusor loci for the trait in question. The progeny of this cross are thenmated back to the superior recurrent parent (A) followed by selection inthe resultant progeny for the desired trait to be transferred from thenon-recurrent parent. After five or more backcross generations withselection for the desired trait, the progeny are heterozygous for locicontrolling the characteristic being transferred, but are like thesuperior parent for most or almost all other loci. The last backcrossgeneration would be selfed to give pure breeding progeny for the traitbeing transferred. When a plant of lettuce variety 81-90 RZ,representative seed of which having been deposited under NCIMB AccessionNo. 42472, is used in backcrossing, offspring retaining the combinationof traits including resistance to downy mildew (Bremia lactucae) racesBl:1-32 and Ca-I to Ca-VIII, resistance to currant-lettuce aphid(Nasonovia ribisnigri), resistance to lettuce mosaic virus (LMV), andresistance to corky root rot (Sphingomonas suberifaciens) are progenywithin the ambit of the invention. Backcrossing methods may be used withthe present invention to improve or introduce a characteristic into aplant of the invention, being a plant of lettuce variety 81-90 RZ. See,e.g., U.S. Pat. No. 7,705,206 (incorporated herein by referenceconsistent with the above INCORPORATION BY REFERENCE section), for ageneral discussion relating to backcrossing.

The invention further involves a method of determining the genotype of aplant of lettuce variety 81-90 RZ, representative seed of which has beendeposited under NCIMB Accession No. 42472, or a first generation progenythereof, which may comprise obtaining a sample of nucleic acids fromsaid plant and detecting in said nucleic acids a plurality ofpolymorphisms. This method may additionally comprise the step of storingthe results of detecting the plurality of polymorphisms on a computerreadable medium. The plurality of polymorphisms are indicative of and/orgive rise to the expression of the morphological and physiologicalcharacteristics of lettuce variety 81-90 RZ.

There are various ways of obtaining genotype data from a nucleic acidsample. Genotype data may be gathered which is specific for certainphenotypic traits (e.g. gene sequences), but also patterns of randomgenetic variation may be obtained to construct a so-called DNAfingerprint. Depending on the technique used a fingerprint may beobtained that is unique for lettuce variety 81-90 RZ. Obtaining a uniqueDNA fingerprint depends on the genetic variation present in a varietyand the sensitivity of the fingerprinting technique. A technique knownin the art to provide a good fingerprint profile is called AFLPfingerprinting technique (See generally U.S. Pat. No. 5,874,215), butthere are many other marker based techniques, such as RFLP (orRestriction fragment length polymorphism), SSLP (or Simple sequencelength polymorphism), RAPD (or Random amplification of polymorphic DNA)VNTR (or Variable number tandem repeat), Microsatellite polymorphism,SSR (or Simple sequence repeat), STR (or Short tandem repeat), SFP (orSingle feature polymorphism) DArT (or Diversity Arrays Technology), RADmarkers (or Restriction site associated DNA markers) (e.g. Baird et al.PloS One Vol. 3 e3376, 2008; Semagn et al. African Journal ofBiotechnology Vol. 5 number 25 pp. 2540-2568, 29 December, 2006).Nowadays, sequence-based methods are utilizing Single NucleotidePolymorphisms (SNPs) that are randomly distributed across genomes, as acommon tool for genotyping (e.g. Elshire et al. PloS One Vol. 6: e19379,2011; Poland et al. PloS One Vol. 7: e32253; Truong et al. PLoS One Vol.7 number 5: e37565, 2012).

With any of the aforementioned genotyping techniques, polymorphisms maybe detected when the genotype and/or sequence of the plant of interestis compared to the genotype and/or sequence of one or more referenceplants. As used herein, the genotype and/or sequence of a referenceplant may be derived from, but is not limited to, any one of thefollowing: parental lines, closely related plant varieties or species,complete genome sequence of a related plant variety or species, or thede novo assembled genome sequence of one or more related plant varietiesor species. For example, it is possible to detect polymorphisms for thecharacteristic downy mildew resistance by comparing the genotype and/orthe sequence of lettuce variety 81-90 RZ with the genotype and/or thesequence of one or more reference plants. The reference plant(s) usedfor comparison in this example may for example be, but is not limitedto, any of the comparison varieties Louxal or Ruby. It is also possiblefor example, to detect polymorphisms for corky root rot resistance bycomparing the genotype and/or the sequence of lettuce variety 81-90 RZwith the genotype and/or the sequence of one or more reference plants.The reference plant(s) used for comparison may for example be, but isnot limited to, any of the comparison varieties Louxal or Ruby.

The polymorphism revealed by these techniques may be used to establishlinks between genotype and phenotype. The polymorphisms may thus be usedto predict or identify certain phenotypic characteristics, individuals,or even species. The polymorphisms are generally called markers. It iscommon practice for the skilled artisan to apply molecular DNAtechniques for generating polymorphisms and creating markers.

The polymorphisms of this invention may be provided in a variety ofmediums to facilitate use, e.g. a database or computer readable medium,which may also contain descriptive annotations in a form that allows askilled artisan to examine or query the polymorphisms and obtain usefulinformation.

As used herein “database” refers to any representation of retrievablecollected data including computer files such as text files, databasefiles, spreadsheet files and image files, printed tabulations andgraphical representations and combinations of digital and image datacollections. In a preferred aspect of the invention, “database” refersto a memory system that may store computer searchable information.

As used herein, “computer readable media” refers to any medium that maybe read and accessed directly by a computer. Such media include, but arenot limited to: magnetic storage media, such as floppy discs, hard disc,storage medium and magnetic tape; optical storage media such as CD-ROM;electrical storage media such as RAM, DRAM, SRAM, SDRAM, ROM; and PROMs(EPROM, EEPROM, Flash EPROM), and hybrids of these categories such asmagnetic/optical storage media. A skilled artisan may readily appreciatehow any of the presently known computer readable mediums may be used tocreate a manufacture which may comprise computer readable medium havingrecorded thereon a polymorphism of the present invention.

As used herein, “recorded” refers to the result of a process for storinginformation in a retrievable database or computer readable medium. Forinstance, a skilled artisan may readily adopt any of the presently knownmethods for recording information on computer readable medium togenerate media which may comprise the polymorphisms of the presentinvention. A variety of data storage structures are available to askilled artisan for creating a computer readable medium where the choiceof the data storage structure will generally be based on the meanschosen to access the stored information. In addition, a variety of dataprocessor programs and formats may be used to store the polymorphisms ofthe present invention on computer readable medium.

The present invention further provides systems, particularlycomputer-based systems, which contain the polymorphisms describedherein. Such systems are designed to identify the polymorphisms of thisinvention. As used herein, “a computer-based system” refers to thehardware, software and memory used to analyze the polymorphisms. Askilled artisan may readily appreciate that any one of the currentlyavailable computer-based system are suitable for use in the presentinvention.

Lettuce leaves are sold in packaged form, including without limitationas pre-packaged lettuce salad or as lettuce heads. Mention is made ofU.S. Pat. No. 5,523,136, incorporated herein by reference consistentwith the above INCORPORATION BY REFERENCE section, which providespackaging film, and packages from such packaging film, including suchpackaging containing leafy produce, and methods for making and usingsuch packaging film and packages, which are suitable for use with thelettuce leaves of the invention. Thus, the invention comprehends the useof and methods for making and using the leaves of the lettuce plant ofthe invention, as well as leaves of lettuce plants derived from theinvention. The invention further relates to a container which maycomprise one or more plants of the invention, or one or more lettuceplants derived from a plant of the invention, in a growth substrate forharvest of leaves from the plant in a domestic environment. This way theconsumer may pick very fresh leaves for use in salads. More generally,the invention includes one or more plants of the invention or one ormore plants derived from lettuce of the invention, wherein the plant isin a ready-to-harvest condition, including with the consumer picking hisown, and further including a container which may comprise one or more ofthese plants.

The invention is further described by the following numbered paragraphs:

1. A lettuce plant of the Batavia type designated 81-90 RZ,representative seed of which having been deposited under NCIMB AccessionNo. 42472, wherein said plant comprises at least the followingcombination of traits including resistance to downy mildew (Bremialactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistance tocurrant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens).

2. The lettuce plant of paragraph 1, wherein the plant has intenseanthocyanin concentration of the mature leaves.

3. A seed of the plant of paragraph 1.

4. A part of the plant of paragraph 1, wherein said part of the plant issuitable for sexual reproduction.

5. The part of the plant of paragraph 4, wherein said part comprises amicrospore, pollen, an ovary, an ovule, an embryo sac or an egg cell.

6. A part of the plant of paragraph 1, wherein said part of the plant issuitable for vegetative reproduction.

7. The part of the plant of paragraph 6, wherein said part comprises acutting, a root, a stem, a cell or a protoplast.

8. A tissue culture of regenerable cells or protoplasts from the lettuceplant of paragraph 1.

9. The tissue culture of paragraph 8, wherein said cells or protoplastsof the tissue culture are derived from a leaf, pollen, an embryo, acotyledon, a hypocotyl, a meristematic cell, a root, a root tip, ananther, a flower, a seed or a stem.

10. A method for producing a progeny plant of a lettuce plant ofparagraph 1, comprising crossing the plant of paragraph 1 with itself orwith another lettuce plant, harvesting the resultant seed, and growingsaid seed.

11. A progeny plant produced by the method of paragraph 10, wherein saidprogeny exhibits a combination of traits including resistance to downymildew (Bremia lactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistanceto currant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens).

12. A progeny plant produced by the method of paragraph 10, wherein saidprogeny plant has all the morphological and physiologicalcharacteristics of the lettuce plant designated 81-90 RZ, representativeseed of which having been deposited under NCIMB Accession No. 42472,wherein said plant comprises at least the following combination oftraits resistance to downy mildew (Bremia lactucae) races Bl:1-32 andCa-I to Ca-VIII, resistance to currant-lettuce aphid (Nasonoviaribisnigri), resistance to lettuce mosaic virus (LMV), and resistance tocorky root rot (Sphingomonas suberifaciens).

13. A progeny plant produced by the method of paragraph 10, wherein saidprogeny plant, representative seed of which having been deposited underNCIMB Accession 42472, is modified in one or more other characteristics.

14. The progeny plant of paragraph 13, wherein the modification iseffected by mutagenesis.

15. The progeny plant of paragraph 13, wherein the modification iseffected by transformation with a transgene.

16. A method of producing a hybrid lettuce seed comprising crossing afirst parent lettuce plant with a second parent lettuce plant andharvesting the resultant hybrid lettuce seed, wherein said first parentlettuce plant or said second parent lettuce plant is the lettuce plantof paragraph 1.

17. A hybrid lettuce seed produced by the method of paragraph 16.

18. A method for producing lettuce leaves as a fresh vegetablecomprising packaging leaves of a plant of paragraph 1.

19. A method for producing lettuce leaves as a processed food comprisingprocessing leaves of a plant of paragraph 1.

20. A container comprising one or more lettuce plants of paragraph 1 forharvest of leaves.

21. A lettuce plant having morphological and/or physiologicalcharacteristics of a lettuce plant, representative seed of which havingbeen deposited under NCIMB Accession No. 42472.

22. A method of introducing a desired trait into a plant of lettucevariety 81-90 RZ comprising: (a) crossing a plant of lettuce variety81-90 RZ, representative seed of which having been deposited under NCIMBAccession No. 42472, with a second lettuce plant that comprises thedesired trait to produce F1 progeny; (b) selecting an F1 progeny thatcomprises the desired trait; (c) crossing the selected F1 progeny with aplant of lettuce variety 81-90 RZ, to produce backcross progeny and (d)selecting backcross progeny comprising the desired trait and thephysiological and morphological characteristic of a plant of lettucevariety 81-90 RZ, when grown in the same environmental conditions.

23. The method of paragraph 22 further comprising (e) repeating steps(c) and (d) one or more times in succession to produce selected fourthor higher backcross progeny that comprise the desired trait and all ofthe physiological and morphological characteristics of a plant oflettuce variety 81-90 RZ, when grown in the same environmentalconditions.

24. A lettuce plant produced by the method of paragraph 22 or 23.

25. A method for producing a seed of a 81-90 RZ-derived lettuce plantcomprising (a) crossing a plant of lettuce variety 81-90 RZ,representative seed of which having been deposited under NCIMB AccessionNo. 42472, with a second lettuce plant, and (b) whereby seed of a 81-90RZ-derived lettuce plant forms.

26. The method of paragraph 25 further comprising (c) crossing a plantgrown from 81-90 RZ-derived lettuce seed with itself or with a secondlettuce plant to yield additional 81-90 RZ-derived lettuce seed, (d)growing the additional 81-90 RZ-derived lettuce seed of step (c) toyield additional 81-90 RZ-derived lettuce plants, and (e) repeating thecrossing and growing of steps (c) and (d) for an additional 3-10generations to generate further 81-90 RZ-derived lettuce plants.

27. The method of paragraph 25 or 26 further comprising selecting atsteps b), d), and e), a 81-90 RZ-derived lettuce plant, exhibiting acombination of traits including resistance to downy mildew (Bremialactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistance tocurrant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens).

28. A seed produced by the method of paragraphs 25, 26, or 27.

29. A method of determining the genotype of a plant of lettuce variety81-90 RZ, representative seed of which has been deposited under NCIMBAccession No. 42472, or a first generation progeny thereof, comprisingobtaining a sample of nucleic acids from said plant and comparing saidnucleic acids to a sample of nucleic acids obtained from a referenceplant, and detecting a plurality of polymorphisms between the twonucleic acid samples, wherein the plurality of polymorphisms areindicative of lettuce variety 81-90 RZ and/or give rise to theexpression of any one or more, or all, of the morphological andphysiological characteristics of lettuce variety 81-90 RZ of paragraph1.

30. The method of paragraph 31 additionally comprising the step ofstoring the results of detecting the plurality of polymorphisms on acomputer readable medium, or transmitting the results of detecting theplurality of polymorphisms.

31. A lettuce plant comprising genetic information for so exhibiting acombination of traits including resistance to downy mildew (Bremialactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistance tocurrant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens), wherein the genetic information is as contained in aplant, representative seed of which having been deposited under NCIMBAccession No. 42472.

32. The lettuce plant of paragraph 33, exhibiting a combination oftraits including resistance to downy mildew (Bremia lactucae) racesBl:1-32 and Ca-I to Ca-VIII, resistance to currant-lettuce aphid(Nasonovia ribisnigri), resistance to lettuce mosaic virus (LMV), andresistance to corky root rot (Sphingomonas suberifaciens), and havingthe genetic information for so exhibiting the combination of traits,wherein the genetic information is as contained in a plant,representative seed of which having been deposited under NCIMB AccessionNo. 42472.

33. The lettuce plant of paragraph 1, which is a plant grown from seedhaving been deposited under NCIMB accession No. 42472.

34. A hybrid lettuce plant grown from the seed of paragraph 17.

35. A method of producing a lettuce cultivar comprising a combination oftraits including resistance to downy mildew (Bremia lactucae) racesBl:1-32 and Ca-I to Ca-VIII, resistance to currant-lettuce aphid(Nasonovia ribisnigri), resistance to lettuce mosaic virus (LMV), andresistance to corky root rot (Sphingomonas suberifaciens), comprising:crossing a mother lettuce plant with a father lettuce plant to produce ahybrid seed; growing said hybrid seed to produce a hybrid plant; selfingsaid hybrid plant to produce F2 progeny seed; growing said F2 progenyseed to produce F2-plants; selecting said F2-plants for exhibitingresistance to downy mildew (Bremia lactucae) races Bl:1-32 and Ca-I toCa-VIII, resistance to currant-lettuce aphid (Nasonovia ribisnigri),resistance to lettuce mosaic virus (LMV), and resistance to corky rootrot (Sphingomonas suberifaciens), and selfing said selected F2-plants toproduce F3 progeny seed; growing said F3 progeny seed to produceF3-plants; selecting F3-plants for exhibiting resistance to downy mildew(Bremia lactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistance tocurrant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens), optionally followed by more selfing and selection steps.

36. A lettuce cultivar produced by the method of paragraph 35.

Having thus described in detail preferred embodiments of the presentinvention, it is to be understood that the invention is not to belimited to particular details set forth in the above description as manyapparent variations thereof are possible without departing from thespirit or scope of the present invention.

What is claimed is:
 1. A lettuce plant of the Batavia type designated81-90 RZ, representative seed of which having been deposited under NCIMBAccession No. 42472, wherein said plant comprises at least the followingcombination of traits including resistance to downy mildew (Bremialactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistance tocurrant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens).
 2. A seed of the plant of claim
 1. 3. A part of theplant of claim 1, wherein said part of the plant is suitable for sexualreproduction.
 4. The part of the plant as claimed in claim 3, whereinsaid part comprises a microspore, pollen, an ovary, an ovule, an embryosac or an egg cell.
 5. A part of the plant of claim 1, wherein said partof the plant is suitable for vegetative reproduction.
 6. The part of theplant as claimed in claim 5, wherein said part comprises a cutting, aroot, a stem, a cell or a protoplast.
 7. A tissue culture of regenerablecells or protoplasts from the lettuce plant of claim
 1. 8. The tissueculture as claimed in claim 7, wherein said cells or protoplasts of thetissue culture are derived from a leaf, pollen, an embryo, a cotyledon,a hypocotyl, a meristematic cell, a root, a root tip, an anther, aflower, a seed or a stem.
 9. A method for producing a progeny plant of alettuce plant of claim 1, comprising crossing the plant of claim 1 withitself or with another lettuce plant, harvesting the resultant seed, andgrowing said seed.
 10. A progeny plant produced by the method of claim9, wherein said progeny plant exhibits a combination of traits includingresistance to downy mildew (Bremia lactucae) races Bl:1-32 and Ca-I toCa-VIII, resistance to currant-lettuce aphid (Nasonovia ribisnigri),resistance to lettuce mosaic virus (LMV), and resistance to corky rootrot (Sphingomonas suberifaciens).
 11. The progeny plant as claimed inclaim 10, wherein said progeny plant is modified in one or more othercharacteristics.
 12. The progeny plant as claimed in claim 11, whereinthe modification is effected by mutagenesis.
 13. The progeny plant asclaimed in claim 11, wherein the modification is effected bytransformation with a transgene.
 14. A progeny plant produced by themethod of claim 9, wherein said progeny plant has all the morphologicaland physiological characteristics of the lettuce plant designated 81-90RZ, representative seed of which having been deposited under NCIMBAccession No.
 42472. 15. A method of producing a hybrid lettuce seedcomprising crossing a first parent lettuce plant with a second parentlettuce plant and harvesting the resultant hybrid lettuce seed, whereinsaid first parent lettuce plant or said second parent lettuce plant isthe lettuce plant of claim
 1. 16. A hybrid lettuce seed produced by themethod of claim
 15. 17. A hybrid lettuce plant grown from the seed asclaimed in claim
 16. 18. A method for producing lettuce leaves as afresh vegetable comprising packaging leaves of a plant of claim
 1. 19. Amethod for producing lettuce leaves as a processed food comprisingprocessing leaves of a plant of claim
 1. 20. A container comprising oneor more lettuce plants of claim 1 for harvest of leaves.
 21. A method ofdetermining the genotype of a plant of lettuce variety 81-90 RZ,representative seed of which has been deposited under NCIMB AccessionNo. 42472, or a first generation progeny thereof, comprising obtaining asample of nucleic acids from said plant and comparing said nucleic acidsto a sample of nucleic acids obtained from a reference plant, anddetecting a plurality of polymorphisms between the two nucleic acidsamples, wherein the plurality of polymorphisms are indicative oflettuce variety 81-90 RZ and/or give rise to the expression of any oneor more, or all, of the morphological and physiological characteristicsof lettuce variety 81-90 RZ as claimed in claim
 1. 22. The method ofclaim 21 additionally comprising the step of storing the results ofdetecting the plurality of polymorphisms on a computer readable medium,or transmitting the results of detecting the plurality of polymorphisms.23. The lettuce plant of claim 1, which is a plant grown from seedhaving been deposited under NCIMB accession No.
 42472. 24. A lettuceplant having resistance to downy mildew (Bremia lactucae) races Bl:1-32and Ca-I to Ca-VIII, resistance to currant-lettuce aphid (Nasonoviaribisnigri), resistance to lettuce mosaic virus (LMV), and resistance tocorky root rot (Sphingomonas suberifaciens) and other morphologicaland/or physiological characteristics of a lettuce plant, representativeseed of which having been deposited under NCIMB Accession No.
 42472. 25.A method of introducing a desired trait into a plant of lettuce variety81-90 RZ comprising: (a) crossing a plant of lettuce variety 81-90 RZ,representative seed of which having been deposited under NCIMB AccessionNo. 42472, with a second lettuce plant that comprises the desired traitto produce F1 progeny; (b) selecting an F1 progeny that comprises thedesired trait; (c) crossing the selected F1 progeny with a plant oflettuce variety 81-90 RZ, to produce backcross progeny and (d) selectingbackcross progeny comprising the desired trait and the physiological andmorphological characteristic of a plant of lettuce variety 81-90 RZ,when grown in the same environmental conditions.
 26. The method of claim25 further comprising (e) repeating steps (c) and (d) one or more timesin succession to produce selected fourth or higher backcross progenythat comprise the desired trait and all of the physiological andmorphological characteristics of a plant of lettuce variety 81-90 RZ,when grown in the same environmental conditions.
 27. A lettuce plantproduced by the method of claim
 25. 28. A method for producing a seed ofa 81-90 RZ-derived lettuce plant comprising (a) crossing a plant oflettuce variety 81-90 RZ, representative seed of which having beendeposited under NCIMB Accession No. 42472, with a second lettuce plant,and (b) whereby seed of a 81-90 RZ-derived lettuce plant forms.
 29. Themethod of claim 28 further comprising (c) crossing a plant grown from81-90 RZ-derived lettuce seed with itself or with a second lettuce plantto yield additional 81-90 RZ-derived lettuce seed, (d) growing theadditional 81-90 RZ-derived lettuce seed of step (c) to yield additional81-90 RZ-derived lettuce plants, and (e) repeating the crossing andgrowing of steps (c) and (d) for an additional 3-10 generations togenerate further 81-90 RZ-derived lettuce plants.
 30. The method ofclaim 29 further comprising selecting at steps b), d), and e), a 81-90RZ-derived lettuce plant, exhibiting a combination of traits includingresistance to downy mildew (Bremia lactucae) races Bl:1-32 and Ca-I toCa-VIII, resistance to currant-lettuce aphid (Nasonovia ribisnigri),resistance to lettuce mosaic virus (LMV), and resistance to corky rootrot (Sphingomonas suberifaciens).
 31. A seed produced by the method ofclaim
 28. 32. A method of producing a lettuce cultivar comprising acombination of traits including resistance to downy mildew (Bremialactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistance tocurrant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens), representative seed of which having been deposited underNCIMB Accession No. 42472, comprising: crossing a mother lettuce plantwith a father lettuce plant to produce a hybrid seed; growing saidhybrid seed to produce a hybrid plant; selfing said hybrid plant toproduce F2 progeny seed; growing said F2 progeny seed to produceF2-plants; selecting said F2-plants for exhibiting resistance to downymildew (Bremia lactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistanceto currant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens), and selfing said selected F2-plants to produce F3progeny seed; growing said F3 progeny seed to produce F3-plants;selecting F3-plants for exhibiting resistance to downy mildew (Bremialactucae) races Bl:1-32 and Ca-I to Ca-VIII, resistance tocurrant-lettuce aphid (Nasonovia ribisnigri), resistance to lettucemosaic virus (LMV), and resistance to corky root rot (Sphingomonassuberifaciens), representative seed of which having been deposited underNCIMB Accession No. 42472, optionally followed by more selfing andselection steps.